Current Organic Chemistry - Volume 18, Issue 8, 2014

General molecular organization of living beings is determined by the flow of energy through the Earth's surface, and is characterized by a limited set of small molecules and polymers, which dominates their chemical constitution. In the 21th century, biology, being previously dominant analytical science has reached a point of being the science of synthesis i.e. synthetic biology. In this perspective, I will argue that the utmost goal of such engineering biology is attempt to change chemical composition of the living cells i.e. to create artificial biodiversity in the frame of carbon-based life chemistry. Thereby, the main obstacle is to find an expedient route to change and expand the fundamental chemistry of life. One of the most promising approaches towards synthetic cell design is the inclusion of amino acid building blocks beyond the canonical 20 (i.e. expanding the genetic code) allowing for alternative reading of the genetic code. Thus, I will briefly elaborate the challenges as well as possible consequences of expanding the genetic code and metabolism of microbial strains for using novel bio-orthogonal chemistries recruited during the engineering process. For example, reprogramming protein biosynthesis with various noncanonical amino acids (ncAAs) will allow the development of industrial microbial strains with enhanced chemical diversity. In consequence, this would enable the creation of safe strains with novel functionalities such as a ‘genetic firewall’, which could potentially be a novel biosafety tool. Such synthetic cells characterized by orthogonal chemistries will have the potential to perform (bio)- chemical transformations currently existing under the exclusive domain of classical synthetic chemistry. In this journey, we will witness the consolidation of Xenobiology (XB), a marriage of chemical synthesis with synthetic biology, to build artificial biological systems for challenging biochemical transformations to address technological problems, while opening the door to a parallel biochemical world. Finally, I will discuss the potential impacts of XB in astrobiology and in the meaning and origin of life.

Chemical synthesis of small molecules is well established and enzyme-mediated chemical synthesis is a growing area, with great potential to be expanded in the future. The recombinant production of proteins has become routine. But molecules of intermediate size remain generally much more difficult to be produced in a sustainable way. Oligopeptides, oligosaccharides and oligonucleotides are three areas orphaned by biotechnology. The industrial manufacturing of all three oligomer types uses almost exclusively chemical synthesis, although conceptually biotechnological methods exist. This review gives an introduction to the market and applications of all three product classes as well as manufacturing options. The future situation could become difficult for all three, but particularly rapidly for peptides, where a number of products are discussed for markets other than the parenteral (injectable) pharmaceuticals. Cost and e-factors of actual chemical methods are by far too high to allow such commercial application, and ecologically and economically sustainable manufacturing methods are urgently needed. The review also shows that similar problems may arise for future pharmaceutical oligonucleotide and oligosaccharide drug products, as aspects of sustainability and cost of goods may significantly limit their broad availability and thereby their potential for commercial success.

Conjugate vaccines have been the most successful way to prevent and control deadly bacterial infections; however their production is complex and tedious. The manufacturing process of these large molecules consists of a biological production and extraction of polysaccharide antigen and protein carrier and its chemical conjugation. Herein a simplified manufacturing process is described where the conjugate vaccine is completely produced in bacterial cells. The polysaccharide antigen and the protein carrier are expressed recombinantly in E. coli cells and linked covalently by an enzymatic process. This novel biological production system leads to improved products and opens up the possibility to develop a wide variety of novel vaccines.

Oligosaccharides and glycoconjugates find application in the food, feed, cosmetic and pharmaceutical industry, which makes glycosylation the target for development of new therapeutic agents, functional food ingredients and other valuable molecules. Unlike other biological molecules, the synthesis of carbohydrates is not template dependent, such that the design of a vast catalog of multifunctional glycostructures is potentially possible. The challenge to achieve regio- and stereo selective structures has made the carbohydrate chemistry one of the most defiant fields of study over the last century, since the synthesis of such a complex molecules is difficult and often impossible to achieve by solely a traditional chemical approach. Enzymatic synthesis however, generally leads to structurally controlled products and overcomes structural complexity. Hence, enzymatic and chemo-enzymatic processes continue gaining attention through the years. Although nature supplies a pool of enzymes able to efficiently catalyze a number of reactions, there are still many transformations requiring redesign at molecular level. Different approaches for the synthesis of valuable glycostructures are covered in this review and their scope and limitations for industrial application are discussed.

Enzyme-catalyzed mono- and oligosaccharide production is a prime example of the potential of multi-enzyme reaction systems, where the regio- and reaction selectivity of these biocatalysts allows the implementation of complex reaction cascades in one vessel where alternative approaches would need lengthy multi-vessel reaction sequences and protection group chemistry. However, this introduces as a new challenge the question of how such multi-enzyme systems can efficiently be assembled in the face of increasing system complexity, which is increasingly addressed by setting up pathways to saccharide production in perforated or intact cells. Put differently, the assembly of such pathways becomes more and more a biosystems engineering activity which needs to take into consideration the need to fine-tune system composition and interactions with the cellular background. This is precisely the domain of synthetic biology, which can be classified as a biosystems engineering activity developing around our increasing capacity to assemble large sets of genes de novo. Here, we argue that biocatalyst construction in saccharide synthesis can broadly benefit from the tools that are being developed in the domain of synthetic biology.

Bioactive peptides are used in diagnostics and as therapeutic agents in a variety of diseases, as well as inhibitors of bacterial growth in food industry. Recent technological advances in delivery and formulation tools have resuscitated the field of peptide therapeutics, resulting in approx. 60 approved peptide drugs and an annual predicted growth rate of the market of approximately 10%. Whilst the majority of peptides are currently produced by chemical synthesis, recombinant peptide production will become more important in the near future and will play a key role in the competition landscape of peptide therapeutics companies. In particular, this is the case for long and complex peptides containing natural amino acids. Although development of a biotechnological process for recombinant production can be time consuming, larger quantities of peptide can be produced and the environmental impact of the generated waste is lower compared to chemical synthesis. However, recombinant peptide expression must overcome several obstacles in order to be cost-effective and competitive with chemical synthesis. The present review focuses on recombinant peptide production in microbial expression platforms, in particular the expression hosts Escherichia coli and yeast and their respective vectors. Strategies which have been successful in solving drawbacks such as degradation of peptides by cellular proteases, solubility and purification issues, toxicity of recombinant peptides for the producer cells and the introduction of posttranslational modifications are described.

Recent advances in coupling antibodies to potent cytotoxic drugs have resulted in stable delivery platforms with improved pharmacokinetics which spares patients from the debilitating systemic toxicity observed with traditional chemotherapy [1]. 5. In 2011 the FDA granted accelerated approval to Seattle Genetics Adcetris® (Bentuximab vedotin) and on Feb. 4th 2013 Health Canada approved the drug which uses a chimeric monoclonal antibody to target CD-30 positive cells. The cytotoxic warhead conjugated to the antibody consists of the potent antimitotic agent monomethyl auristatin E. This is the first new therapy approved since 1977 for treatment of Hodgkin’s lymphoma and it is the first therapy to receive approval against anaplastic large cell lymphoma. Roche-Genentech received approval to offer Kadcyla® (Trastuzumab emtansine) for the treatment of patients with HER2-positive metastatic breast cancer in Feb. 2013. Results from the pivotal EMILIA trial demonstrated significantly improved overall survival compared to current standard of care which uses capecitabine and lapatinib. Kadcyla is the Herceptin antibody conjugated to the maytansine derivative DM1. Manufacturing these highly potent biopharmaceuticals presents a series of unique engineering and chemistry challenges which has resulted in several contract manufacturers constructing facilities to address the unique engineering and chemistry challenges associated with this promising class of therapeutic drugs. By leveraging experience in biopharmaceuticals and potent small molecule drug process development this class of promising new therapies can be reliably and safely manufactured to meet clinical and commercial demands. This article will investigate the experience of Lonza as it synergistically pooled internal resources to address this promising class of therapeutics.

Compared with the conventional luminescent molecules, whose fluorescence is quenched once they aggregate, molecules with aggregation-induced emission (AIE) properties exhibit significantly enhanced emission in solid state or aggregates due to their unique molecular structures and stacking modes, showing potential applications in optoelectronic devices, biochemical sensors and bioimaging. Especially, AIE fluorophores recently have drawn much attention as an emerging sensory material. Taking advantage of AIE effects, fluorescence signals can be boosted by AIE dyes in presence of analytes, which offers a promising platform for sensor with intense fluorescence signal. In this review, we highlight the recent advances in the development of AIE fluorescent probes that are classified into various categories, including tetraphenylethenes, siloles, 9,10-distyrylanthracene, cyanostilbene and systems with extended π-conjugations to those even without typical AIE chromophores. The fluorescent behaviors of these probes toward different analyte are discussed.

Isoprenoids constitute the largest class of natural products with greater than 55,000 identified members. They play essential roles in maintaining proper cellular function leading to maintenance of human health, plant defense mechanisms against predators, and are often exploited for their beneficial properties in the pharmaceutical and nutraceutical industries. Most impressively, all known isoprenoids are derived from one of two C5-precursors, isopentenyl diphosphate (IPP) or dimethylallyl diphosphate (DMAPP). In order to study the enzyme transformations leading to the extensive structural diversity found within this class of compounds there must be access to the substrates. Sometimes, intermediates within a biological pathway can be isolated and used directly to study enzyme/pathway function. However, the primary route to most of the isoprenoid intermediates is through chemical catalysis. As such, this review provides the first exhaustive examination of synthetic routes to isoprenoid and isoprenoid precursors with particular emphasis on the syntheses of intermediates found as part of the 2C-methylerythritol 4-phosphate (MEP) pathway. In addition, representative syntheses are presented for the monoterpenes (C10), sesquiterpenes (C15, diterpenes (C20), triterpenes (C30) and tetraterpenes (C40). Finally, in some instances, the synthetic routes to substrate analogs found both within the MEP pathway and downstream isoprenoids are examined.